SUMMARY Intestinal absorption of dietary fats is important for the development of obesity and its complications. The long- term goal is to understand the role of the integrin family of cell surface matrix receptors in regulating intestinal nutrient absorption. The overall objective of this application is to elucidate the role of the ?v?3 and ?v?5 integrins in regulating small intestinal lipid homeostasis. The central hypothesis is that in response to ingestion of dietary fat, enterocytes secrete the integrin ligand Mfge8. Mfge8 then binds enterocyte ?v?3 and ?v?5 integrins activating a signaling pathway that increases the absorption of dietary fat coupled with synthesis of triglyceride (TG) in the endoplasmic reticulum and hydrolysis of TG in cytoplasmic lipid droplets. This hypothesis is based on data demonstrating that mice deficient in Mfge8 or the ?v?3 and ?v?5 integrins develop steatorrhea and in response to a dietary fat challenge absorb less fat, accumulate excess cytoplasmic lipid droplets and secrete less chylomicrons. The rationale for these studies is that delineating the role of the Mfge8-integrin axis in intestinal lipid absorption is necessary to understand the mechanisms by which enterocytes regulate lipoprotein production. This hypothesis will be tested through 3 specific aims: 1) Determining how nutrients induce enterocyte production of Mfge8 transcript and protein and the physiological role of the Mfge8 and the ?v?3 and ?v?5 integrins in absorption of dietary fat; 2) Determining the role of the ?v?3 and ?v?5 integrins in regulating hydrolysis of TG in enterocyte cytoplasmic lipid droplets; and 3) Determining the therapeutic potential of targeting the ?v?3 and ?v?5 integrins for the treatment of obesity. Aim 1 will examine transcriptional regulation of Mfge8 in differentiated Caco-2 cells and primary enterocytes, secretion of Mfge8 into the intestinal lumen in response to different nutrients, and identification of the cellular source of Mfge8 required for fat absorption through use of transgenic mice with enterocyte specific deletion of Mfge8 or Mfge8 deficient mice with enterocyte specific expression of Mfge8. Aim 2 will examine the mechanisms by which Mfge8 and the ?v?3 and ?v?5 integrins regulate hydrolysis of TG from lipid droplets using in vivo studies with integrin and Mfge8 deficient mice and in vitro studies with differentiated Caco-2 cells. Studies will determine how the Mfge8-integrin axis regulates subcellular localization and phosphorylation of lipolytic mediators. Aim 3 will test the therapeutic efficacy of oral inhibition of the ?v?3 and ?v?5 integrins in inducing weight loss in obese mice coupled with development of novel compounds with optimized potency and selectivity for blocking this pathway. The proposed research is innovative, in the applicant's opinion, because it identifies a mechanism for linking uptake of ingested dietary fats with hydrolysis of TG from cytoplasmic lipid droplets for chylomicron production. The proposed research is significant because it will expand the understanding of how the intestine absorbs and processes dietary fat for export. This knowledge has the potential to inform the development of therapeutics that can reduce obesity and postprandial lipemia.